Collagen is a fibrous protein that form fibrils having a very high tensile strength and that has been found in most multicellular organisms. Collagen serves to hold cells and tissues together and to direct the development of mature tissue. Collagen is the major fibrous protein in skin, cartilage, bone, tendon, blood vessels and teeth.
There are many types of collagen which differ from each other to meet the requirements of various tissues. Some examples of types of collagen are as follows: type one [.alpha.1(I)].sub.2 .alpha.2 which is found in skin, tendon, bone and cornea; type two [.alpha.1(II)].sub.3 which is found in cartilage intervertebral disc, and the vitreous body; type three [.alpha.1(III)].sub.3 which can be found in skin and the cardiovascular system; type four [.alpha.1(IV)].sub.2 .alpha.2(IV) which can be found in basement membrane; type five [.alpha.1(V)].sub.2 .alpha.2(V) and .alpha.1(V).alpha.2(V).alpha.3(V) which is found in the placenta and cornea. Examples of newly identified forms of collagen include: type seven (VII) which is found in anchoring fibrils beneath many epithelia; and types nine (IX), ten (X) and eleven (XI), which are minor constituents of cartilage.
The chemical characterization of native collagen was difficult since its low solubility made isolation of collagen a tedious task. Eventually, it was discovered that collagen from tissues of young animals was not as extensively crosslinked as that of mature tissues and thus was more amenable to extraction. For example, the basic structural unit of type I collagen, tropo-collagen, can be extracted in intact form from some young, collagen-containing animal tissues.
The extraction of collagen from collagen-containing tissues is of interest since collagen can be repolymerized to form fibrils that make a valuable gel supporting cell growth, or a matrix suitable for shaping into prosthetic structures, synthetic skin or membranes. Presently, salt solutions are being used to extract the salt soluble fractions of collagen which is a small fraction with minimal crosslinking. Dilute acid solutions are also being used (i.e., acetic, citric, formic, and hydrochloric acids) to extract collagen from young rapidly growing tissues. The acid-soluble fraction is slightly greater than the salt-soluble fraction; however, in some instances, the extraction solutions may be more dangerous to handle due to their acidic pH. See Gross et al., Proc. Natl. Acad. Sci U.S.A., 41, pp 1-7 (1955) and Davison et al, Conn. Tiss. Res., 1, pp 205-216, (1972). Insoluble collagen is also digested with pepsin to solubilize an additional fraction.
Nimni et al., Biochemical and Biophysical Research Communications, 25, No. 4, pp 434-440 (1966) teach the extraction of collagen from the skin of rats using penicillamine or cysteamine in a NaCl solution. Both penicillamine and cysteamine are very expensive solvents which have not been widely used. The extraction method of Nimni et al. uses penicillamine or cysteamine to disrupt the crosslinks prior to extracting with a sodium chloride solution. Nimni et al. does not teach the regeneration of collagen after extraction.